A Composition for treatment of a bacterial infection of the digestive tract

ABSTRACT

A composition for treatment of bacterial infections of the digestive tract is disclosed which comprises a lytic enzyme composition specific for the infecting bacteria, and a carrier for delivering said lytic enzyme. The bacteria to be treated is selected from the group consisting of Listeria, Salmonella,  E. coli , Campylobacter, and combinations thereof. The carrier for delivering at least one lytic enzyme to the digestive tract is selected from the group consisting of suppository enemas, syrups, or enteric coated pills.

[0001] The following application is a continuation-in-part of Ser. No.09/497,495 filed Apr. 18, 2000 which is a continuation of Ser. No.09/395,636 filed Sep. 14, 2000, now U.S. Pat. No. 6,056,954 which is acontinuation-in-part of U.S. patent application Ser. No. 08/962,523,filed Oct. 31, 1997, now U.S. Pat. No. 5,997,862.

DESCRIPTION Background of the Invention

[0002] 1. Field of the Invention

[0003] The present invention discloses a method and composition for thetreatment of bacterial infections by the use of a lysing enzyme blendedwith an appropriate carrier suitable for the treatment of the infection.

[0004] 2. Description of the Prior Art

[0005] In the past, antibiotics have been used to treat variousinfections. The work of Selman Waksman in the introduction andproduction of Streptomycetes, Dr. Fleming's discovery of penicillin, arewell known as well as the work of numerous others in the field ofantibiotics. Over the years, there have been additions and chemicalmodifications to the “basic” antibiotics in attempts to make them morepowerful, or to treat people allergic to these antibiotics.

[0006] Others have found new uses for these antibiotics. U.S. Pat. No.5,260,292 (Robinson et al.) discloses the topical treatment of acne withaminopenicillins. The method and composition for topically treating acneand acneiform dermal disorders includes applying an amount of anantibiotic selected from the group consisting of ampicillin,amoxicillin, other aminopenicillins, and cephalosporins, and derivativesand analogs thereof, effective to treat the acne and acneiform dermaldisorders. U.S. Pat. No. 5,409,917 (Robinson et al.) discloses thetopical treatment of acne with ceplhalosporins.

[0007] However, as more antibiotics have been prescribed or used at anever increasing rate for a variety of illnesses, increasing numbers ofbacteria have developed a resistance to antibiotics. Larger doses ofstronger antibiotics are now being used to treat ever more resistantstrains of bacteria. Multiple antibiotic resistant bacteria haveconsequently developed. The use of more antibiotics and the number ofbacteria showing resistance has led to increasing the amount of timethat the antibiotics need to be used. Broad, non-specific antibiotics,some of which have detrimental effects on the patient, are now beingused more frequently. Also, antibiotics do not easily penetrate mucuslinings.

[0008] Additionally, the number of people allergic to antibioticsappears to be increasing.

[0009] Consequently, other efforts have been sought to first identifyand then kill bacteria.

[0010] Attempts have been made to treat bacterial diseases with by theuse of bacteriophages. U.S. Pat. No. 5,688,501 (Merril, et al.)discloses a method for treating an infectious disease caused by bacteriain an animal with lytic or non-lytic bacteriophages that are specificfor particular bacteria.

[0011] U.S. Pat. No. 4,957,686 (Norris) discloses a procedure ofimproved dental hygiene which comprises introducing into the mouthbacteriophages parasitic to bacteria which possess the property ofreadily adhering to the salivary pellicle.

[0012] It is to be noted that the direct introduction of bacteriophagesinto an animal to prevent or fight diseases has certain drawbacks.Specifically, the bacteria must be in the right growth phase for thephage to attach. Both the bacteria and the and the phage have to be inthe correct and synchronized growth cycles. Additionally, there must bethe right number of phages to attach to the bacteria; if there are toomany or too few phages, there will either be no attachment or noproduction of the lysing enzyme. The phage must also be active enough.The phages are also inhibited by many things including bacterial debrisfrom the organism it is going to attack. Further complicating the directuse of bacteriophage to treat bacterial infections is the possibility ofimmunological reactions, rendering the phage non-functional.

[0013] Consequently, others have explored the use of other safer andmore effective means to treat and prevent bacterial infections.

[0014] U.S. Pat. No. 5,604,109 (Fischetti et al.) relates to the rapiddetection of Group A streptococci in clinical specimens, through theenzymatic digestion by a semi-purified Group C streptococcal phageassociated lysin enzyme. The lytic enzyme of this patent is used in U.S.Pat. No. 5,997,862 (Fischetti, et. al.), U.S. Pat. No. 5,985,271,(Fischetti et al.) and U.S. Pat. No. 6,017,528 (Fischetti et al.) whichdisclose the use of an oral delivery mode, such as a candy, chewing gum,lozenge, troche, tablet, a powder, an aerosol, a liquid or a liquidspray, containing a lysin enzyme produced by group C streptococcalbacteria infected with a C1 bacteriophage for the prophylactic andtherapeutic treatment of Streptococcal A throat infections, commonlyknown as strep throat.

[0015] U.S. Pat. No. 6,056,955 (Fischetti et al.) discloses the topicaltreatment of streptococcal infections.

SUMMARY OF THE INVENTION

[0016] The method for obtaining and purifying the lytic enzyme producedby a bacteria infected with the bacteriophage is known in the art. Somerecent evidence suggests that the phage enzyme that lyses thestreptococcus organism may actually be a bacterial enzyme that is usedto construct the cell wall and the phage. While replicating in thebacterium, a phage gene product may cause the upregulation orderepression of bacterial enzyme for the purpose of releasing thebacteriophage. These bacterial enzymes may be tightly regulated by thebacterial cell and are used by the bacteria for the construction andassembly of the cell wall.

[0017] The use of these lytic enzymes for the prophylactic andtherapeutic treatment of bacterial diseases, however, has not beenexplored, except by the inventors of the present invention.Consequently, the present invention discloses the extraction and use ofa variety of bacterial phage associated lytic enzymes for the treatmentof a wide variety of illnesses caused by bacterial infections.

[0018] The use of phage associated lytic enzymes produced by theinfection of a bacteria with a bacteria specific phage has numerousadvantages for the treatment of diseases. As the phage are targeted forspecific bacteria, the lytic enzymes do not interfere with normal flora.Also, lytic phages primarily attack cell wall structures which are notaffected by plasmid variation. The actions of the lytic enzymes are fastand do not depend on bacterial growth.

[0019] Lytic enzymes can be directed to the mucosal lining, where, inresidence, they will be able to kill colonizing bacteria.

[0020] It is an object of the invention to use phage associated enzymesto prophylactically and therapeutically treat bacterial diseases.

[0021] The invention (which incorporates U.S. Pat. No. 5,604,109 in itsentirety by reference) uses an enzyme produced by the bacterial organismafter being infected with a particular bacteriophage as either aprophylactic treatment for preventing those who have been exposed toothers who have the symptoms of an infection from getting sick, or as atherapeutic treatment for those who have already become ill from theinfection. The present invention is based upon the discovery that phagelytic enzymes specific for bacteria infected with a specific phage caneffectively and efficiently break down the cell wall of the bacterium inquestion. At the same time, the semipurified enzyme is lacking inproteolytic enzymatic activity and therefore non-destructive tomammalian proteins and tissues when present during the digestion of thebacterial cell wall. The same general technique used to produce andpurify the lysin enzyme in U.S. Pat. No. 5,604,109 may be used tomanufacture other lytic enzymes produced by bacteria infected with abacteriophage specific for that bacteria. Depending on the bacteria,there may be variations in the growth media and conditions.

[0022] In one embodiment of the invention, the prophylactic andtherapeutic treatment of a variety of illnesses caused by Streptococcalpneumoniae, Streptococcus fasciae, and Hemophilus influenza aredisclosed. In another embodiment of the invention, gram negativebacterial infections caused by Listeria, Salmonella, E. coli, andCampylobacter, are treated by the use of lytic enzymes. These and otherbacteria, which can infect the digestive system, can be treated byincorporating the lytic enzymes in suppository enemas, in syrups, or inother carriers to get directly to the site of the infection(s).

[0023] In another embodiment of the invention, lytic enzymes areincorporated into bandages to prevent or treat infections of burns andwounds. In yet another embodiment of the invention, the lytic enzymes ofphage associated with Staphylococcus or Pseudomonas are incorporatedinto bandages to prevent or treat infections of burns and wounds.

[0024] Vaginal infections caused by Group B Streptococcus can causepremature birth and subsequent complications resulting in neonatalsepsis. Lysin incorporated into tampons specific for group B strep wouldprevent infection of the neonate during birth without disturbing normalvaginal flora so that women would not be overcome by yeast infection asa result of antibiotic therapy.

[0025] In another embodiment of the invention, eye drops containinglytic enzymes of Hemophilus, Pseudomonas, and/or Staphylococcus can beused to directly treat eye infections. Treatment with lytic enzymes arefaster and more expedient than with antibiotics.

[0026] In yet another embodiment of the invention the phage associatedlytic enzyme is put into a carrier which is placed in an inhaler totreat or prevent the spread of diseases localized in the mucus lining ofthe oral cavity and lungs. Specific lytic enzymes for tuberculosis havebeen isolated and can be used.

[0027] In another embodiment of the invention the lytic enzyme isadministered in the form of a candy, chewing gum, lozenge, troche,tablet, a powder, an aerosol, a liquid, a liquid spray, or toothpastefor the prevention or treatment of bacterial infections associated withupper respiratory tract illnesses.

[0028] In another embodiment of the invention, species specific lyticenzymes can be used in the treatment of bacterial infections associatedwith topical or dermatological infections, administered in the form of atopical ointment or cream. In another embodiment of the invention, thelytic enzyme would be administered in an aqueous form. In yet anotherembodiment of the invention, lysostaphin, the enzyme which lysesStaphylococcus aureus, can be included in the therapeutic agent. In afurther embodiment of the invention, conventional antibiotics may beincluded in the therapeutic agent with the lytic enzyme, and with orwithout the presence of lysostaphin. More than one lytic enzyme may alsobe included in the prophylactic or therapeutic agent.

DETAILED DESCRIPTION OF THE INVENTION

[0029] The method for treating bacterial infections comprises treatingthe infection with a therapeutic agent comprising an effective amount ofat least one lytic enzyme produced by a bacteria infected with abacteriophage specific for the bacteria. The lytic enzyme is preferablyin an environment having a pH which allows for activity of said lyticenzyme.

[0030] The lytic enzyme can be used for the treatment or prevention ofHemophilus influenza, Pseudomonas, Streptococcus pneumoniae,Streptococcus fasciae, Streptococcus group B, Listeria, Salmonella, E.coli, Campylobacter, and other bacteria, and any combination thereof.

[0031] For example, if there is a bacterial infection of the upperrespiratory tract, the infection can be prophylactically ortherapeutically treated with a composition comprising an effectiveamount of at least one lytic enzyme produced by a bacteria beinginfected with a bacteriophage specific for that bacteria, and a carrierfor delivering the lytic enzyme to a mouth, throat, or nasal passage. Itis preferred that the lytic enzyme is in an environment having a pHwhich allows for activity of the lytic enzyme. If an individual has beenexposed to someone with the upper respiratory disorder, the lytic enzymewill reside in the mucosal lining and prevent any colonization of theinfecting bacteria.

[0032] Two examples of bacteria which infect the upper respiratorysystem are Streptococcus pneumoniae and Hemophilus influenzae. In recentyears, there has been an increase in the number of people, particularlychildren and the elderly, that are infected or are carriers ofpenicillin resistant Streptococcus pneumoniae and Hemophilus. Whilethese bacteria are normally harmless residents of the host, they areopportunistic organisms that are able to cause infections when theresistance of the host has been compromised. By eliminating or reducingthe number of these organisms in the upper respiratory tract, there willbe a commensurate reduction in the number of infections by thesebacteria.

[0033] Infection of the Hemophilus bacteria by Bacteriophage HP1 (amember of the P2-like phage family with strong similarities tocoliphages P2 and 186, and some similarity to the retronphage Ec67)produces a lytic enzyme capable of lysing the bacteria. The lytic enzymefor Streptococcus pneumoniae, previously identified as anN-acetyl-muramoyl-L-alanine amidase, is produced by the infectingStreptococcus pneumoniae with the Pal bacteriophage. The therapeuticagent can contain either or both of the lytic enzymes produced by thesetwo bacteria, and may contain other lytic enzymes for other bacteria.The composition which may be used for the prophylactic and therapeutictreatment of a strep infection includes the lysin enzyme and a means ofapplication, (such as a carrier system or an oral delivery mode), to themucosal lining of the oral and nasal cavity, such that the enzyme is putin the carrier system or oral delivery mode to reach the mucosa lining.Another infection which can be treated prophylactically Streptococcusgroup A, which can produce what is commonly known as “strep” throat.When group C Streptococci are infected with a C1 bacteriophage, a lysinenzyme is produced specific for the lysing of Streptococcus group A.

[0034] Prior to, or at the time the lysin enzyme is put in the carriersystem or oral delivery mode, it is preferred that the enzyme be in astabilizing buffer environment for maintaining a pH range between about4.0 and about 9.0, more preferably between about 5.5 and about 7.5 andmost preferably at about 6.1.

[0035] The stabilizing buffer should allow for the optimum activity ofthe lysin enzyme. The buffer may be a reducing reagent, such asdithiothreitol. The stabilizing buffer may also be or include a metalchelating reagent, such as ethylenediaminetetracetic acid disodium salt,or it may also contain a phosphate or citrate-phosphate buffer.

[0036] Means of application include, but are not limited to direct,indirect, carrier and special means or any combination of means. Directapplication of the lytic enzyme may be by nasal sprays, nasal drops,nasal ointments, nasal washes, nasal injections, nasal packings,bronchial sprays and inhalers, or indirectly through use of throatlozenges, or through use of mouthwashes or gargles, or through the useof ointments applied to the nasal nares, the bridge of the nose, or theface or any combination of these and similar methods of application. Theforms in which the lysin enzyme may be administered include but are notlimited to lozenges, troches, candies, injectants, chewing gums,tablets, powders, sprays, liquids, ointments, and aerosols.

[0037] The lozenge, tablet, or gum into which the lytic enzyme is addedmay contain sugar, corn syrup, a variety of dyes, non-sugar sweeteners,flavorings, any binders, or combinations thereof. Similarly, any gumbased products may contain acacia, carnauba wax, citric acid, cornstarch, food colorings, flavorings, non-sugar sweeteners, gelatin,glucose, glycerin, gum base, shellac, sodium saccharin, sugar, water,white wax, cellulose, other binders, and combinations thereof.

[0038] Lozenges may further contain sucrose, corn starch, acacia, gumtragacanth, anethole, linseed, oleoresin, mineral oil, and cellulose,other binders, and combinations thereof. In another embodiment of theinvention, sugar substitutes are used in place of dextrose, sucrose, orother sugars.

[0039] The enzyme may also be placed in a nasal spray, wherein the nasalspray is the carrier. The nasal spray can be a long acting or timedrelease spray, and can be manufactured by means well known in the art.An inhalant may also be used, so that the phage enzyme may reach furtherdown into the bronchial tract, including into the lungs.

[0040] Any of the carriers for the lytic enzyme may be manufactured byconventional means. However, it is preferred that any mouthwash orsimilar type products not contain alcohol to prevent denaturing of theenzyme. Similarly, when the lytic enzyme is being placed in a coughdrop, gum, candy or lozenge during the manufacturing process, suchplacement should be made prior to the hardening of the lozenge or candybut after the cough drop or candy has cooled somewhat, to avoid heatdenaturation of the enzyme.

[0041] The enzyme may be added to these substances in a liquid form orin a lyophilized state, whereupon it will be solubilized when it meetsbody fluids such as saliva. The enzyme may also be in a micelle orliposome.

[0042] The effective dosage rates or amounts of the lytic enzyme totreat the infection will depend in part on whether the lytic will beused therapeutically or prophylactically, the duration of exposure ofthe recipient to the infectious bacteria, the size and weight of theindividual, etc. The duration for use of the composition containing theenzyme also depends on whether the use is for prophylactic purposes,wherein the use may be hourly, daily or weekly, for a short time period,or whether the use will be for therapeutic purposes wherein a moreintensive regimen of the use of the composition may be needed, such thatusage may last for hours, days or weeks, and/or on a daily basis, or attimed intervals during the day. Any dosage form employed should providefor a minimum number of units for a minimum amount of time. Theconcentration of the active units of enzyme believed to provide for aneffective amount or dosage of enzyme may be in the range of about 100units/ml to about 100,000 units/ml of fluid in the wet or dampenvironment of the nasal and oral passages, and possibly in the range ofabout 100 units/ml to about 10,000 units/ml. More specifically, timeexposure to the active enzyme units may influence the desiredconcentration of active enzyme units per ml. It should be noted thatcarriers that are classified as “long” or “slow” release carriers (suchas, for example, certain nasal sprays or lozenges) could possess orprovide a lower concentration of active (enzyme) units per ml, but overa longer period of time, whereas a “short” or “fast” release carrier(such as, for example, a gargle) could possess or provide a highconcentration of active (enzyme) units per ml, but over a shorter periodof time. The amount of active units per ml and the duration of time ofexposure depends on the nature of infection, whether treatment is to beprophylactic or therapeutic, and other variables.

[0043] While this treatment may be used in any mammalian species, thepreferred use of this product is for a human.

[0044] This composition and method may also be used for the treatment ofStreptococcus A infections of the respiratory tract. When using thiscomposition for a Streptococcus A infection, the lysin phage enzymeshould be used for the prophylactic prevention of Streptococcusinfections. Similarly, in another embodiment of the invention, thismethod may be used for the therapeutic and, preferably, the prophylactictreatment of tuberculosis. In a preferred embodiment of the invention,the phage associated lysing enzyme for Mycobacteria tuberculosis isplaced in a carrier in an inhaler. The carrier may be sterile water or awater base, or any other carrier used in an inhaler for dispersing drugsinto the bronchial tract. The phage associated lytic enzyme specific fortuberculosis is subject to the same conditions as the phage associatedlytic enzyme for other lytic enzymes. Specifically, prior to, or at thetime the enzyme is put in the carrier system or oral delivery mode, itis preferred that the enzyme be in a stabilizing buffer environment formaintaining a pH range between about 4.0 and about 9.0.

[0045] The stabilizing buffer should allow for the optimum activity ofthe lytic enzyme. The buffer may be a reducing reagent, such asdithiothreitol. The stabilizing buffer may also be or include a metalchelating reagent, such as ethylenediaminetetracetic acid disodium salt,or it may also contain a phosphate or citrate-phosphate buffer.

[0046] For the prophylactic and therapeutic treatment of tuberculosis,the phage associated lytic enzyme associated with tuberculosis may alsobe applied by direct, indirect, carriers and special means or anycombination of means. Direct application of the lytic enzyme may be bynasal sprays, nasal drops, nasal ointments, nasal washes, nasalinjections, nasal packings, bronchial sprays and inhalers, or indirectlythrough use of throat lozenges, or through use of mouthwashes orgargles, or through the use of ointments applied to the nasal nares, thebridge of the nose, or the face or any combination of these and similarmethods of application. The forms in which the lytic enzyme may beadministered include but are not limited to lozenges, troches, candies,injectants, chewing gums, tablets, powders, sprays, liquids, ointments,and aerosols. For the therapeutic treatment of tuberculosis, thebronchial sprays and aerosols are most beneficial, as these carriers, ormeans of distributing the composition, allow the lytic enzyme to reachthe bronchial tubes and the lungs. An appropriate transport carrier maybe attached to the enzyme to transport the enzyme across the cellmembrane to the site of the bacteria.

[0047] The lozenge, tablet, or gum into which the lytic enzyme is addedmay contain sugar, corn syrup, a variety of dyes, non-sugar sweeteners,flavorings, any binders, or combinations thereof. Similarly, any gumbased products may contain acacia, carnauba wax, citric acid, cornstarch, food colorings, flavorings, non-sugar sweeteners, gelatin,glucose, glycerin, gum base, shellac, sodium saccharin, sugar, water,white wax, cellulose, other binders, and combinations thereof.

[0048] Lozenges may further contain sucrose, corn starch, acacia, gumtragacanth, anethole, linseed, oleoresin, mineral oil, and cellulose,other binders, and combinations thereof. In another embodiment of theinvention, sugar substitutes are used in place of dextrose, sucrose, orother sugars. However, to tackle bacterial infections in the lung, theuse of an inhaler carrier the lytic enzyme in a carrier is preferred.

[0049] Another use of a lytic enzyme is for the treatment of bacterialinfections of the digestive tract. The method for treating a bacterialinfection of the digestive tract comprises treating the bacterialinfection with composition comprising an effective amount of at leastone lytic enzyme produced by a bacteria infected with a bacteriophagespecific for the bacteria, and a carrier for delivering said lyticenzyme to the digestive tract. In a preferred embodiment of theinvention, the bacterial infections being treated are being caused bygram negative bacteria selected from the group consisting of Listeria,Salmonella, E. coli, and Campylobacter. However, this method andcomposition will effectively treat other bacteria, when the appropriatelytic enzyme is used.

[0050] In a preferred embodiment of the invention, the carrier isselected from the group consisting of suppository enemas, syrups, orenteric coated pills. These proposed carriers can be made byconventional methods. However, the only difference in their manufactureis that the enzyme being placed in the carrier must not be allowed todenature. To that end, the enzyme should be incorporated into a carrierwhich does not contain alcohol, and which has been cooled to atemperature that will not cause the denaturing of the enzyme. The enzymemay be incorporated in a lyophilized state, or may be incorporated in aliposome before being placed in the suppository, syrup or enteric coatedpill.

[0051] The enzyme placed in the composition or carrier should be in anenvironment having a pH which allows for activity of the lytic enzyme.To this end, the pH of the composition is preferably kept in a range ofbetween about 2 and about 11, more preferably in a range of betweenabout between about 4.0 and about 9.0, and even more preferably at a pHrange of between about 5.5 and about 7.5. As described above with theother lytic enzyme, the pH can be moderated by the use of a buffer. Thebuffer may contain a reducing agent, and more specificallydithiothreitol. The buffer may also be a a metal chelating reagent, suchas ethylenediaminetetracetic disodium salt or the buffer may contain acitrate-phosphate buffer. As with all compositions described in thispatent, the composition may further include a bactericidal orbacteriostatic agent as a preservative.

[0052] The lytic enzyme is preferably present in a concentration ofabout 100 to about 500,000 active enzyme units per milliliter of fluidin the wet environment of the gastrointestinal tract, preferably about100 to about 100,000 active enzyme units per milliliter of fluid, andpreferably present in a concentration of about 100 to about 10,000active enzyme units per milliliter of fluid in the wet environment ofthe gastrointestinal tract.

[0053] The suppository is known in the art, and is made of glycerin,fatty acids, and similar type substances that dissolve at bodytemperature. As the suppository dissolves, the phage associated lyticenzyme will be released.

[0054] Another composition and use of the lytic enzyme is for thetherapeutic or prophylactic treatment of bacterial infections of burnsand wounds of the skin. The composition comprises an effective amount ofat least one lytic enzyme produced by a bacteria infected with abacteriophage specific for the bacteria and a carrier for delivering atleast one lytic enzyme to the wounded skin. The mode of application forthe lytic enzyme includes a number of different types and combinationsof carriers which include, but are not limited to an aqueous liquid, analcohol base liquid, a water soluble gel, a lotion, an ointment, anonaqueous liquid base, a mineral oil base, a blend of mineral oil andpetrolatum, lanolin, liposomes, protein carriers such as serum albuminor gelatin, powdered cellulose carmel, and combinations thereof. A modeof delivery of the carrier containing the therapeutic agent includes butis not limited to a smear, spray, a time-release patch, a liquidabsorbed wipe, and combinations thereof. The lytic enzyme may be appliedto a bandage either directly or in one of the other carriers. Thebandages may be sold damp or dry, wherein the enzyme is in a lyophilizedform on the bandage. This method of application is most effective forthe treatment of burns.

[0055] The carriers of the compositions of the present invention maycomprise semi-solid and gel-like vehicles that include a polymerthickener, water, preservatives, active surfactants or emulsifiers,antioxidants, sun screens, and a solvent or mixed solvent system. U.S.Pat. No. 5,863,560 (Osborne) discusses a number of different carriercombinations which can aid in the exposure of the skin to a medicament.

[0056] Polymer thickeners that may be used include those known to oneskilled in the art, such as hydrophilic and hydroalcoholic gellingagents frequently used in the cosmetic and pharmaceutical industries.Preferably, the hydrophilic or hydroalcoholic gelling agent comprises“CARBOPOL.RTM.” (B. F. Goodrich, Cleveland, Ohio), “HYPAN.RTM.”(Kingston Technologies, Dayton, N.J.), “NATROSOL.RTM.” (Aqualon,Wilmington, Del.), “KLUCEL.RTM.” (Aqualon, Wilmington, Del.), or“STABILEZE.RTM.” (ISP Technologies, Wayne, N.J.). Preferably, thegelling agent comprises between about 0.2% to about 4% by weight of thecomposition. More particularly, the preferred compositional weightpercent range for “CARBOPOL.RTM.” is between about 0.5% to about 2%,while the preferred weight percent range for “NATROSOL.RTM.” and“KLUCEL.RTM.” is between about 0.5% to about 4%. The preferredcompositional weight percent range for both “HYPAN.RTM.” and“STABILEZE.RTM.” is between about 0.5% to about 4%. “CARBOPOL.RTM.” isone of numerous cross-linked acrylic acid polymers that are given thegeneral adopted name carbomer. These polymers dissolve in water and forma clear or slightly hazy gel upon neutralization with a caustic materialsuch as sodium hydroxide, potassium hydroxide, triethanolamine, or otheramine bases. “KLUCEL.RTM.” is a cellulose polymer that is dispersed inwater and forms a uniform gel upon complete hydration. Other preferredgelling polymers include hydroxyethylcellulose, cellulose gum, MVE/MAdecadiene crosspolymer, PVM/MA copolymer, or a combination thereof.

[0057] Preservatives may also be used in this invention and preferablycomprise about 0.05% to 0.5% by weight of the total composition. The useof preservatives assures that if the product is microbiallycontaminated, the formulation will prevent or diminish microorganismgrowth. Some preservatives useful in this invention includemethylparaben, propylparaben, butylparaben, chloroxylenol, sodiumbenzoate, DMDM Hydantoin, 3-Iodo-2-Propylbutyl carbamate, potassiumsorbate, chlorhexidine digluconate, or a combination thereof.

[0058] Titanium dioxide may be used as a sunscreen to serve asprophylaxis against photosensitization. Alternative sun screens includemethyl cinnamate. Moreover, BHA may be used as an antioxidant, as wellas to protect ethoxydiglycol and/or dapsone from discoloration due tooxidation. An alternate antioxidant is BHT.

[0059] Pharmaceuticals for use in all embodiments of the inventioninclude antimicrobial agents, anti-inflammatory agents, antiviralagents, local anesthetic agents, corticosteroids, destructive therapyagents, antifungals, and antiandrogens. In the treatment of acne, activepharmaceuticals that may be used include antimicrobial agents,especially those having anti-inflammatory properties such as dapsone,erythromycin, minocycline, tetracycline, clindamycin, and otherantimicrobials. The preferred weight percentages for the antimicrobialsare 0.5% to 10%.

[0060] Local anesthetics include tetracaine, tetracaine hydrochloride,lidocaine, lidocaine hydrochloride, dyclonine, dyclonine hydrochloride,dimethisoquin hydrochloride, dibucaine, dibucaine hydrochloride,butambenpicrate, and pramoxine hydrochloride. A preferred concentrationfor local anesthetics is about 0.025% to 5% by weight of the totalcomposition. Anesthetics such as benzocaine may also be used at apreferred concentration of about 2% to 25% by weight.

[0061] Corticosteroids that may be used include betamethasonedipropionate, fluocinolone actinide, betamethasone valerate,triamcinolone actinide, clobetasol propionate, desoximetasone,diflorasone diacetate, amcinonide, flurandrenolide, hydrocortisonevalerate, hydrocortisone butyrate, and desonide are recommended atconcentrations of about 0.01% to 1.0% by weight. Preferredconcentrations for corticosteroids such as hydrocortisone ormethylprednisolone acetate are from about 0.2% to about 5.0% by weight.

[0062] Destructive therapy agents such as salicylic acid or lactic acidmay also be used. A concentration of about 2% to about 40% by weight ispreferred. Cantharidin is preferably utilized in a concentration ofabout 5% to about 30% by weight. Typical antifungals that may be used inthis invention and their preferred weight concentrations include:oxiconazole nitrate (0.1% to 5.0%), ciclopirox olamine (0.1% to 5.0%),ketoconazole (0.1% to 5.0%), miconazole nitrate (0.1% to 5.0%), andbutoconazole nitrate (0.1% to 5.0%). For the topical treatment ofseborrheic dermatitis, hirsutism, acne, and alopecia, the activepharmaceutical may include an antiandrogen such as flutamide orfinasteride in preferred weight percentages of about 0.5% to 10%.

[0063] Typically, treatments using a combination of drugs includeantibiotics in combination with local anesthetics such as polymycin Bsulfate and neomycin sulfate in combination with tetracaine for topicalantibiotic gels to provide prophylaxis against infection and relief ofpain. Another example is the use of minoxidil in combination with acorticosteroid such as betamethasone diproprionate for the treatment ofalopecia ereata. The combination of an anti-inflammatory such ascortisone with an antifungal such as ketoconazole for the treatment oftinea infections is also an example.

[0064] In one embodiment, the invention comprises a dermatologicalcomposition having about 0.5% to 10% carbomer and about 0.5% to 10% of apharmaceutical that exists in both a dissolved state and a microparticulate state. The dissolved pharmaceutical has the capacity tocross the stratum corneum, whereas the micro particulate pharmaceuticaldoes not. Addition of an amine base, potassium, hydroxide solution, orsodium hydroxide solution completes the formation of the gel. Moreparticularly, the pharmaceutical may include dapsone, an antimicrobialagent having anti-inflammatory properties. A preferred ratio of microparticulate to dissolved dapsone is five or less.

[0065] In another embodiment, the invention comprises about 1% carbomer,about 80-90% water, about 10% ethoxydiglycol, about 0.2% methylparaben,about 0.3% to 3.0% dapsone including both micro particulate dapsone anddissolved dapsone, and about 2% caustic material. More particularly, thecarbomer may include “CARBOPOL.RTM. 980” and the caustic material mayinclude sodium hydroxide solution.

[0066] In a preferred embodiment, the composition comprises dapsone andethoxydiglycol, which allows for an optimized ratio of micro particulatedrug to dissolved drug. This ratio determines the amount of drugdelivered, compared to the amount of drug retained in or above thestratum corneum to function in the supracorneum domain. The system ofdapsone and ethoxydiglycol may include purified water combined with“CARBOPOL.RTM.” gelling polymer, methylparaben, propylparaben, titaniumdioxide, BHA, and a caustic material to neutralize the “CARBOPOL.RTM.”

[0067] Any of the carriers for the lytic enzyme may be manufactured byconventional means. However, if alcohol is used in the carrier, theenzyme should be in a micelle, liposome, or a “reverse” liposome, toprevent denaturing of the enzyme. Similarly, when the lytic enzyme isbeing placed in the carrier, and the carrier is, or has been heated,such placement should be made after the carrier has cooled somewhat, toavoid heat denaturation of the enzyme. In a preferred embodiment of theinvention, the carrier is sterile.

[0068] The enzyme may be added to these substances in a liquid form orin a lyophilized state, whereupon it will be solubilized when it meets aliquid body.

[0069] The effective dosage rates or amounts of the lytic enzyme totreat the infection, and the duration of treatment will depend in parton the seriousness of the infection, the duration of exposure of therecipient to the infectious bacteria, the number of square centimetersof skin or tissue which are infected, the depth of the infection, theseriousness of the infection, and a variety of a number of othervariables. The composition may be applied anywhere from once to severaltimes a day, and may be applied for a short or long term period. Theusage may last for days or weeks. Any dosage form employed shouldprovide for a minimum number of units for a minimum amount of time. Theconcentration of the active units of enzyme believed to provide for aneffective amount or dosage of enzyme may be in the range of about 100units/ml to about 500,000 units/ml of composition, preferably in therange of about 1000 units/ml to about 100,000 units/ml, and mostpreferably from about 10,000 to 100,000 units/ml. The amount of activeunits per ml and the duration of time of exposure depends on the natureof infection, and the amount of contact the carrier allows the lyticenzyme to have. It is to be remembered that the enzyme works best whenin a fluid environment. Hence, effectiveness of the enzyme is in partrelated to the amount of moisture trapped by the carrier. In anotherpreferred embodiment, a mild surfactant in an amount effective topotentiate the therapeutic effect of the lytic enzyme. Suitable mildsurfactants include, inter alia, esters of polyoxyethylene sorbitan andfatty acids (Tween series), octylphenoxypolyethoxy ethanol (Triton-Xseries), n-Octyl-.beta.-D-glucopyranoside,n-Octyl-.beta.-D-thioglucopyranoside, nDecyl-.beta.-D-glucopyranoside,n-Dodecyl-.beta.-D-glucopyranoside, and biologically occurringsurfactants, e.g., fatty acids, glycerides, monoglycerides, deoxycholateand esters of deoxycholate.

[0070] In order to accelerate treatment of the infection, thetherapeutic agent may further include at least one complementary agentwhich can also potentiate the bactericidal activity of the lytic enzyme.The complementary agent can be penicillin, synthetic penicillinsbacitracin, methicillin, cephalosporin, polymyxin, cefaclor. Cefadroxil,cefamandole nafate, cefazolin, cefixime, cefmetazole, cefonioid,cefoperazone, ceforanide, cefotanme, cefotaxime, cefotetan, cefoxitin,cefpodoxime proxetil, ceftazidime, ceftizoxime, ceftriaxone, cefriaxonemoxalactam, cefuroxime, cephalexin, cephalosporin C, cephalosporin Csodium salt, cephalothin, cephalothin sodium salt, cephapirin,cephradine, cefuroximeaxetil, dihydratecephalothin, moxalactam,loracarbef.mafate, chelating agents and any combinations thereof inamounts which are effective to synergistically enhance the therapeuticeffect of the lytic enzyme.

[0071] Additionally, the therapeutic agent may further comprise theenzyme lysostaphin for the treatment of any Staphylococcus aureusbacteria. Mucolytic peptides, such as lysostaphin, have been suggestedto be efficacious in the treatment of S. aureus infections of humans(Schaffner et al., Yale J. Biol. & Med., 39:230 (1967) and bovinemastitis caused by S. aureus (Sears et al., J. Dairy Science, 71 (Suppl.1): 244(1988)). Lysostaphin, a gene product of Staphylococcus simulans,exerts a bacteriostatic and bactericidal effect upon S. aureus byenzymatically degrading the polyglycine crosslinks of the cell wall(Browder et al., Res. Comm., 19: 393-400 (1965)). U.S. Pat. No.3,278,378 describes fermentation methods for producing lysostaphin fromculture media of S. staphylolyticus, later renamed S. simulans. Othermethods for producing lysostaphin are further described in U.S. Pat.Nos. 3,398,056 and 3,594,284. The gene for lysostaphin has subsequentlybeen cloned and sequenced (Recsei et al., Proc. Natl. Acad. Sci. USA,84: 1127-1131 (1987)). The recombinant mucolytic bactericidal protein,such as r-lysostaphin, can potentially circumvent problems associatedwith current antibiotic therapy because of its targeted specificity, lowtoxicity and possible reduction of biologically active residues.Furthermore, lysostaphin is also active against non-dividing cells,while most antibiotics require actively dividing cells to mediate theireffects (Dixon et al., Yale J. Biology and Medicine, 41: 62-68 (1968)).Lysostaphin, in combination with the lysin enzyme, can be used in thepresence or absence of the listed antibiotics. There is a degree ofadded importance in using both lysostaphin and the lysin enzyme in thesame therapeutic agent. Frequently, when a body has a bacterialinfection, the infection by one genus of bacteria weakens the body orchanges the bacterial flora of the body, allowing other potentiallypathogenic bacteria to infect the body. One of the bacteria thatsometimes co-infects a body is Staphylococcus aureus. Many strains ofStaphylococcus aureus produce penicillinase, such that Staphylococcus,Streptococcus, and other gram positive bacterial strains will not bekilled by standard antibiotics. Consequently, the use of the lysin andlysostaphin, possibly in combination with antibiotics, can serve as themost rapid and effective treatment of bacterial infections. In yetanother preferred embodiment, the invention may include mutanolysin, andlysozyme

[0072] In preferred embodiments of the invention, the lytic enzymes forPseudomonas, Staphylococcus, and Streptococcus, jointly or individually,may be incorporated into the carrier, or into a bandage to be used onburn patients, or in a solution or cream carrier.

[0073] Yet another use of lytic enzymes is for the prophylactic ortherapeutic treatment of vaginal infections. This treatment comprisestreating the vaginal infection with an effective amount of at least onelytic enzyme produced by a bacteria being infected with a bacteriophagespecific for that bacteria, wherein that lytic enzyme is incorporated ina carrier to be placed in a vagina. The preferred carrier is a tampon,or vaginal douche. A pad may also be used as a carrier, although it isnot as effective. While any number of bacteria could be treated usingthis composition and method, it is believed that the most optimum use ofthis treatment composition and method would be for the treatment of anE. coli and Streptococcus B infection. Vaginal infections caused byGroup B Streptococcus can cause neonatal meningitis resulting in braindamage and premature death. Lytic enzyme incorporated into tamponspecific for group B Strep would eliminate the group B organisms withoutdisturbing normal flora so that woman would not be overcome by yeastinfection post antibiotic therapy. The use of the lytic enzyme in thevagina would best provide a prophylactic effect, although therapeuticuse would also be advisable.

[0074] To produce a pad or tampon containing the enzyme, the lyticenzyme can be applied in a solution to the tampon, and allowed to dry.The lytic enzyme may be incorporated into the pad or tampon by any othermeans known in the art, including lyophilization, spraying, etc. Thetampons and pads may also be kept slightly moist, and in a sealedwrapper until ready for use. In that case, bactericide andbacteriostatic compounds and inhibitors should be present in the tamponsand pads. The method to be used for incorporating the lytic enzyme intothe tampon or pad can be one of the methods known in the art forincorporating a pharmaceutical product. In another embodiment of theinvention, the lytic enzyme is incorporated into a vaginal suppository.The vaginal suppository into which the lytic enzyme is beingincorporated may be a standard vaginal suppository, comprised ofglyceride, alginate, starch, other standard binders and any combinationsthereof.

[0075] When using a tampon as the carrier, it is best to insert thetampon in the vagina and leave it in for up to 12 hours to distributethe enzyme vaginally.

[0076] As with other lytic enzymes, it is preferable that the pH be keptin a range of about 4.0 and about 9.0 even more preferably at a pH rangeof between about 5.5 and about 7.5. As described above with the otherlytic enzyme, the pH can be moderated by the use of a buffer. The buffermay contain a reducing agent, and more specifically dithiothreitol. Thebuffer may also contain a metal chelating reagent, such asethylenediaminetetracetic disodium salt or the buffer may be acitrate-phosphate buffer. As with all compositions described in thispatent, the composition may, further include a bactericidal orbacteriostatic agent as a preservative.

[0077] The lytic enzyme is preferably present in a concentration ofabout 100 to about 500,000 active enzyme units per milliliter of fluidin the wet environment of the vaginal tract, preferably about 100 toabout 100,000 active enzyme units per milliliter of fluid, andpreferably present in a concentration of about 100 to about 10,000active enzyme units per milliliter of fluid in the wet environment ofthe vaginal tract.

[0078] Another use of the invention is for the prophylactic andtherapeutic treatment of eye infections. The method of treatmentcomprises administering eye drops which comprise an effective amount ofat least one lytic enzyme produced by the bacteria being infected with abacteriophage specific for the bacteria and a carrier capable of beingsafely applied to an eye, with the carrier containing the lytic enzyme.In a preferred embodiment of the invention, the bacteria being treatedis Hemophilus or Staphylococcus The eye drops are in the form of anisotonic solution. The pH of the solution should be adjusted so thatthere is no irritation of the eye, which in turn would lead to possiblyinfection by other organisms, and possibly to damage to the eye. Whilethe pH range should be in the same range as for other lytic enzymes, themost optimal pH will be in the range of from 6.0 to 7.5. Similarly,buffers of the sort described above for the other lytic enzymes shouldalso be used. Other antibiotics which are suitable for use in eye dropsmay be added to the composition containing the lytic enzymes.Bactericides and bacteriostatic compounds may also be added.

[0079] It is to be remembered that all of the enzymes can be used forprophylactic and therapeutic treatments of the bacteria for which theenzymes are specific.

[0080] It is also to be remembered that a carrier may have more than onelytic enzyme. For instance, A throat lozenge may comprise just a lysinenzyme (which lyses the Streptococcus A strain causing “strep” throat,or it may also include the lytic enzymes for Hemophilus. Similarly, thecarrier for treating burns and wounds, or infections of the skin, maycontain just one lytic enzyme, or a combination of lytic enzymes, forthe treatment of Pseudomonas, Streptococcus, Staphylococcus, or anyother of a number of bacteria.

[0081] Lytic enzymes can also be used to fight dental caries.Specifically, a lytic enzyme specific for Streptococcus mutans may beincorporated in a toothpaste or oral wash. Similarly, this lytic enzymemay also be incorporated into a chewing gum or lozenge. Any othercarrier can be used that allows for the exposure of the mouth, gums, andteeth to the lytic enzyme.

[0082] The lytic enzyme may also be incorporated in a lyophilized ordried form in tooth powder. If the lytic enzyme is to be used in an oralwash, it is preferred that the oral wash not contain any alcohol, so asto not denature the enzyme. The enzyme can also be in a liposome whenmixed in with the toothpaste or oral wash. The concentrations of theenzyme units per ml of toothpaste or mouth wash can be in the range offrom about 100 units/ml to about 500,000 units/ml of composition,preferably in the range of about 1000 units/ml to about 100,000units/ml, and most preferably from about 10,000 to 100,000 units/ml. ThepH of the toothpaste or oral wash should be in a range that allows forthe optimum performance of the enzyme, while not causing any discomfortto the user of the toothpaste or oral wash.

[0083] Many modifications and variations of the present invention arepossible in light of the above teachings. It is, therefore, to beunderstood within the scope of the appended claims the invention may beprotected otherwise than as specifically described.

What we claim is: 1) A composition for treatment of bacterial infectionsof the digestive tract, comprising an effective amount of at least onelytic enzyme produced by said bacteria being infected with abacteriophage specific for said bacteria, and a carrier for deliveringsaid at least one lytic enzyme to the digestive tract. 2) Thecomposition according to claim 1, wherein said bacteria to be treated isselected from the group consisting of Listeria, Salmonella, E. coli,Campylobacter, and combinations thereof. 3) The composition according toclaim 1, wherein said carrier for delivering said at least one lyticenzyme to the digestive tract is selected from the group consisting ofsuppository enemas, syrups, or enteric coated pills. 4) The compositionaccording to claim 1, wherein said composition further comprises abuffer that maintains pH of the composition at a range between about 4.0and 9.0. 5) The composition according to claim 4, wherein the buffermaintains the pH of the composition at the range between 5.5 and 7.5. 6)The composition according to claim 4, wherein said buffer comprises areducing reagent. 7) The composition according to claim 6, wherein saidreducing reagent is dithiothreitol. 8) The composition according toclaim 4, wherein said buffer comprises a metal chelating reagent. 9) Thecomposition according to claim 8, wherein said metal chelating reagentis ethylenediaminetetracetic disodium salt. 10) The compositionaccording to claim 4, wherein said buffer is a citrate-phosphate buffer.11) The composition according to claim 1, further comprising abactericidal or bacteriostatic agent as a preservative. 12) Thecomposition according to claim 1, wherein said at least one lytic enzymeis lyophilized. 13). The composition according claim 1, wherein said atleast one lytic enzyme is present in a concentration of about 100 toabout 100,000 active enzyme units per milliliter of fluid in the wetenvironment of the digestive tract 14). The composition according toclaim 13, wherein said at least one lytic enzyme is present in aconcentration of about 100 to about 10,000 active enzyme units permilliliter of fluid in the wet environment of the digestive tract.